Search

Objective: To evaluate whether M2 macrophage-derived exosomes protect against MI/R injury and reveal the protective mechanism of exosomes [Kourembanas 2015]. Methods: I/R model injury was induced by temporary left anterior descending coronary artery occlusion in Sprague-Dawley (SD) rats, macrophages isolated from bone marrow-derived macrophages (BMDMs) were induced to M2 polarization, and H9C2 cells subjected to hypoxia/reperfusion (H/R) were used to establish an in vitro model. I/R-induced rats and H/R-induced H9C2 cells were treated with M2-exos in vivo and in vitro, respectively. Masson staining was performed to observe myocardial fibrosis in rats. Immunohistochemical (IHC) staining of myocardial tissues showed the expression of NLRP3 inflammasome activation and pyrolysis. Exosomes derived from IL-4-treated macrophages (M2-exos) were detected by transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA) and western bolt. Western bolt was performed to determine the protein level, including NLRP3, pro-caspase-1, cleaved caspase-1, pro-IL-1β, cleaved IL-1β, gasdermin D (GSDMD), and N-terminus of gasdermin D (GSDMD-N). Results: Activity of NLRP3 inflammasome and existence of pyroptosis in the rats subjected to MI/R were significantly higher than those in the control (P < 0.05). Moreover, we confirmed the accumulation of ROS during I/R injury in cardiomyocytes. M2-exos protected against I/R injury and reduced activity of NLRP3 inflammasome and existence of pyroptosis, accompanied with attenuating oxidative stress. In vitro studies showed similar effects, H9c2 cells co-cultured with M2-exos could attenuated H/R-induced cell injury, while M2-exos suppressed the expression of NLRP3 inflammasome and pyroptosis (P < 0.05).

Cardiovascular diseases, including congenital and acquired cardiovascular diseases, impose a severe burden on healthcare systems worldwide. Although bone marrow-derived stem cells (BMSCs) therapy can be an effective therapeutic strategy for the heart disease, relatively low abundance, difficult accessibility, and small tissue volume hinder the clinical usefulness. Adipose tissue-derived stem cells (ADSCs) show similar potential with BMSCs to differentiate into lineages and tissues, such as smooth muscle cells, endothelial cells, and adipocytes, with attractiveness of obtaining adipose tissue easily and repeatedly, and a simple separation procedure. We briefly summarize the current understanding of the cardiomyocytes differentiated from ADSCs

Hypoxia-induced pulmonary arterial hypertension (PAH) is characterized by prostacyclin (PGI2 ) disorder, which manifests in the same manner as in monocrotaline (MCT)-induced PAH. Endogenous PGI2 inhibitor coupling factor 6 (CF6) is involved in MCT-induced PAH. This study aimed to explore the presence or absence of a correlation between hypoxia-induced PAH and CF6.This study was conducted between January 2019 and June 2020. A total of 135 male Wistar rats (aged 8 weeks and weighing 200-250 g) were randomly divided into five groups: (A) control, (B) 1 week of hypoxia, (C) 2 weeks of hypoxia, (D) 3 weeks of hypoxia, and (E) 4 weeks of hypoxia. CF6 expression in both lung tissue and blood samples from the lung vasculature and tail vein was measured by western blotting, immunohistochemistry, reverse transcription polymerase chain reaction, and enzyme-linked immunosorbent assay.Hemodynamic and morphological changes in hypoxia-induced rats indicated PAH development. The results showed the presence of a correlation between the mRNA and protein levels of CF6 in lung tissue, activity of mitochondrial ATP synthase, and hypoxia time, and there was a significant increment in the group exposed to hypoxia for 4 weeks compared to the control group. The decrement expression of ATPase inhibitory factor 1 (IF 1) mRNA was consistent with the outcomes of ATP synthase activity in lung tissue in the 4 weeks of hypoxia group compared with the control group. However, the levels of CF6 and ATP synthase activity did not differ between blood samples from the lung vasculature and tail vein.: In hypoxia-induced PAH, CF6 showed downregulated expression in lung tissue, but not in pulmonary vasculature and circulation. Therefore, we speculated that CF6 and ATP synthase may play important roles in hypoxia-induced PAH.

Purpose Pulmonary arterial hypertension (PAH) is a disease that the pulmonary artery is abnormally elevated. P2Y12 is an adenosine diphosphate (ADP) receptor and it act as the target of thienopyridine antiplatelet drugs by controlling vascular remodeling. Inhibition of P2Y12 receptor in the process of PAH was explored in this study. Methods The PAH model was established in Sprague-Dawley rats by single subcutaneous injection of 60 mg/kg monocrotaline (MCT). The ticagrelor solution (a selective P2Y12R inhibitor) was intraperitoneally injected into rats at a dose of 14 mg/kg from the time of MCT injection to day 28. Results In the lung tissues of PAH rats, the marked P2Y12R was detected. Treatment with ticagrelor greatly decreased P2Y12R level and efficiently abolished the upregulation of α-SMA as demonstrated by Western blot and RT-PCR. The wall thickness and occlusion score of the pulmonary arterioles showed that blockade of P2Y12R could relieve lung remodeling caused by PAH. The haemodynamic changes at 4 weeks determined that P2Y12R inhibition affected RV pressure and right heart hypertrophy. Conclusions P2Y12R might be involved in the pathogenesis of PAH. Blockade of P2Y12R has potential in treating PAH.

Myocardial infarction (MI) is one of the leading causes of high mortality worldwide. Long non-coding RNA myocardial infarction associated transcript (MIAT) and mitochondrial coupling factor 6 (CF6) aggravate MI. This study aimed to elucidate whether miR-203 interacted with MIAT and CF6 in MI. Results revealed that MIAT and CF6 expressions were upregulated and that miR-203 was downregulated in mouse myocardial tissues after MI, as well as in hypoxic mouse cardiomyocytes. The overexpression of MIAT in mouse cardiomyocytes raised CF6 expression, whereas the knockdown of MIAT had the opposite effect. Mechanistically, the luciferase reporter and RNA pull-down assays corroborated the binding between miR-203 and CF6 3'UTR and between miR-203 and MIAT. The simultaneous overexpression of miR-203 and MIAT restored the reduction of CF6 caused by miR-203 overexpression alone, and the overexpression of miR-203 diminished the percentage of infarct area and the apoptosis of cardiomyocytes in vivo. Our findings corroborate that overexpressing miR-203 alleviates MI via interacting with MIAT and CF6.

Sympathetic activation after myocardial infarction (MI) leads to ventricular arrhythmias (VAs), which can result in sudden cardiac death (SCD). The toll-like receptor 4 (TLR4)/myeloid differentiation primary response 88 (MyD88)/nuclear factor-kappa B (NF-kB) axis within the hypothalamic paraventricular nucleus (PVN), a cardiac-neural sympathetic nerve centre, plays an important role in causing VAs. An MI rat model and a PVN-TLR4 knockdown model were constructed. The levels of protein were detected by Western blotting and immunofluorescence, and localizations were visualized by multiple immunofluorescence staining. Central and peripheral sympathetic activation was visualized by immunohistochemistry for c-fos protein, renal sympathetic nerve activity (RSNA) measurement, heart rate variability (HRV) analysis and norepinephrine (NE) level detection in serum and myocardial tissue measured by ELISA. The arrhythmia scores were measured by programmed electrical stimulation (PES), and cardiac function was detected by the pressure-volume loop (P-V loop). The levels of TLR4 and MyD88 and the nuclear translocation of NF-kB within the PVN were increased after MI, while sympathetic activation and arrhythmia scores were increased and cardiac function was decreased. However, inhibition of TLR4 significantly reversed these conditions. PVN-mediated sympathetic activation via the TLR4/MyD88/NF-kB axis ultimately leads to the development of VAs after MI.

Ventricular arrhythmias (VAs) triggers by sympathetic nerve hyperactivity contribute to sudden cardiac death in myocardial infarction (MI) patients. Microglia-mediated inflammation in the paraventricular nucleus (PVN) is involved in sympathetic hyperactivity after MI. N6-methyladenosine (m

Sudden cardiac death caused by ventricular arrhythmias (VAs) is the main cause of high mortality in patients with myocardial infarction (MI). Sympathetic neural remodeling caused by inflammation after MI is closely associated with the occurrence of VAs. METTL3, the earliest identified mA rat MI model was established via left coronary artery ligation. The expression of METTL3, TRAF6, NOX2, and NF-κB increased at 3 days and remained elevated at 7 days after MI, as determined via Western blotting. METTL3 was primarily present in macrophages, as determined via immunofluorescence. Intramyocardial injection of lentivirus carrying METTL3-shRNA inhibited METTL3 expression in vivo. Methylated immunoprecipitation-qPCR determined the METTL3 knockdown inhibited the mDownregulation of METTL3 expression attenuated the excessive sympathetic neural remodeling induced by MI, further reducing the incidence of VAs and improving cardiac function. This was partly associated with the inhibition of the TRAF6/NF-κB pathway and ROS production.

Background: Pulmonary arterial hypertension (PAH) is a disease that the pulmonary artery is abnormally elevated. P2Y12 is an adenosine diphosphate (ADP) receptor and it act as the target of thienopyridine antiplatelet drugs by controlling vascular remodeling. Inhibition of P2Y12 receptor in the process of PAH was explored in this study.Methods: The PAH model was established in Sprague-Dawley rats by single subcutaneous injection of 60 mg/kg monocrotaline (MCT). The ticagrelor solution (a selective P2Y12R inhibitor) was intraperitoneally injected into rats at a dose of 14 mg/kg from the time of MCT injection to day 28.Results: In the lung tissues of PAH rats, the marked P2Y12R was detected. Treatment with ticagrelor greatly decreased P2Y12R level and efficiently abolished the upregulation of α-SMA as demonstrated by Western blot and RT-PCR. The wall thickness and occlusion score of the pulmonary arterioles showed that blockade of P2Y12R could relieve lung remodeling caused by PAH. The haemodynamic changes at 4 weeks determined that P2Y12R inhibition affected RV pressure and right heart hypertrophy.Conclusions: P2Y12R might be involved in the pathogenesis of PAH. Blockade of P2Y12R has potential in treating PAH.

Cerebral malaria (CM) is a severe complication with brain vascular hyperpermeability. Claudin-5 is the major component of tight junctions. To investigate the expression of claudin-5 in CM, we established a murine experimental cerebral malaria (ECM) model and an in vitro model by treating murine brain endothelial cells (bEnd3) with plasma from ECM mice. Expression of claudin-5 and the ETS transcription factor Erg was reduced in the brain endothelium of ECM mice. In bEnd3 cells exposed to ECM plasma, decreased expression of claudin-5 and Erg, and increased permeability were observed. Silencing of Erg significantly reduced Cldn5 expression. ChIP assays indicated that Erg binds to the -813 ETS motif of the murine Cldn5 gene promoter, and the binding is decreased by treatment with ECM plasma.

Background The hypothalamic paraventricular nucleus (PVN) is the center of the regulation of autonomic nervous system functions and cardiovascular activity. Phosphoinositide-3 kinase (PI3K)-AKT pathway in PVN contributes to mediate sympathetic nerve activity and is activated in spontaneously hypertensive rats. Overactivation of the sympathetic output was considered as an important mechanism of the arrhythmias. In the present study, we aimed to explore whether targeted regulation of sympathetic activity in PVN could reduce the peripheral sympathoexcitatory and attenuate the ventricular arrhythmias (VAs) in myocardial infarction (MI) rats via PI3K-AKT pathway. Methods A stainless steel gauge guide cannula was stereotaxically implanted into the PVN, and 7 days later, rats were randomly divided into the following 4 groups: group A, control + dimethyl sulfoxide (DMSO); group B, control + LY294002; group C, MI surgery + DMSO; and group D, MI surgery + LY294002. Studies were conducted seven days post-MI. Myocardial function, infarct size, inducible VAs by programmed electrical stimulation, renal sympathetic nerve activity (RSNA), and protein level of PI3K and AKT were measured. Results MI increased the protein ratios of p-PI3K-to-total-PI3K and p-AKT-to-total-AKT in PVN but can be reduced by LY294002 treatment. Inhibition of sympathetic nerve activity in PVN led to a reversion in plasma norepinephrine, RSNA and inducible VAs in MI rats. Conclusions PI3K-AKT pathway in the PVN was a main mechanism in regulating sympathetic activity and arrhythmias in MI rats. Targeted inhibition of sympathetic activity in PVN may be a potential treatment for the VAs via PI3K-AKT pathway.

Background Cardiac sympathetic hyperinnervation after myocardial infarction (MI) is associated with a high incidence of lethal arrhythmia. Erythropoietin-producing hepatoma interactor B2 (EphrinB2), a diffusible axonal chemorepellent that can induce growth cone collapse and axon repulsion of several neuronal populations, is crucial in neurodevelopment during disease development and progression. However, whether EphrinB2 could inhibit cardiac sympathetic hyperinnervation after MI remains unclear. Methods and results A rat model of MI was developed by left anterior descending coronary artery ligation. EphrinB2 expression was markedly increased in the infarcted border at 3 days after MI. Downregulation of EphrinB2 by intramyocardial injection of lentivirus carrying EphrinB2-shRNA significantly increased sympathetic hyperinnervation along with downregulated RhoA expression. In contrast, injection of EphrinB2-overexpressing lentivirus markedly upregulated EphrinB2, concomitant with inhibition of sympathetic sprouting and upregulated RhoA expression, accompanied by decreased incidence of ventricular arrhythmias (VAs). However, co-administering EphrinB2-overexpressing lentivirus and Fasudil (Rho kinase inhibitor) nearly abolished the inhibition of nerve sprouting effect. Additionally, EphrinB2 expression did not affect nerve growth factor level in the infarcted heart. Conclusions Overexpression of EphrinB2 may ameliorate MI-induced sympathetic hyperinnervation and further reduce the incidence of VAs, at least in part by activating RhoA-mediated axonal retraction.

Myocardial infarction (MI) is one of the leading causes of high mortality worldwide. Long non-coding RNA myocardial infarction associated transcript (MIAT) and mitochondrial coupling factor 6 (CF6) aggravate MI. This study aimed to elucidate whether miR-203 interacted with MIAT and CF6 in MI. Results revealed that MIAT and CF6 expressions were upregulated and that miR-203 was downregulated in mouse myocardial tissues after MI, as well as in hypoxic mouse cardiomyocytes. The overexpression of MIAT in mouse cardiomyocytes raised CF6 expression, whereas the knockdown of MIAT had the opposite effect. Mechanistically, the luciferase reporter and RNA pull-down assays corroborated the binding between miR-203 and CF6 3'UTR and between miR-203 and MIAT. The simultaneous overexpression of miR-203 and MIAT restored the reduction of CF6 caused by miR-203 overexpression alone, and the overexpression of miR-203 diminished the percentage of infarct area and the apoptosis of cardiomyocytes in vivo. Our findings corroborate that overexpressing miR-203 alleviates MI via interacting with MIAT and CF6.

Malignant ventricular arrhythmias (VAs) following myocardial infarction (MI) is a lethal complication resulting from sympathetic nerve hyperactivity. Numerous evidence have shown that inflammation within the paraventricular nucleus (PVN) participates in sympathetic hyperactivity. Our aim was to explore the role of Macrophage‐inducible C‐type lectin (Mincle) within the PVN in augmenting sympathetic activity following MI,and whether NOD‐like receptor family pyrin domain‐containing 3 (NLRP3) inflammasome/IL‐1β axis is involved in this activity. MI was induced by coronary artery ligation. Mincle expression localized in microglia within the PVN was markedly increased at 24 hours post‐MI together with sympathetic hyperactivity, as indicated by measurement of the renal sympathetic nerve activity (RSNA) and norepinephrine (NE) concentration. Mincle‐specific siRNA was administrated locally to the PVN, which consequently decreased microglial activation and sympathetic nerve activity. The MI rats exhibited a higher arrhythmia score after programmed electric stimulation than that treated with Mincle siRNA, suggesting that the inhibition of Mincle attenuated foetal ventricular arrhythmias post‐MI. The underlying mechanism of Mincle in sympathetic hyperactivity was investigated in lipopolysaccharide (LPS)‐primed naïve rats. Recombinant Sin3A‐associated protein 130kD (rSAP130), an endogenous ligand for Mincle, induced high levels of NLRP3 and mature IL‐1β protein. PVN‐targeted injection of NLRP3 siRNA or IL‐1β antagonist gevokizumab attenuated sympathetic hyperactivity. Together, the data indicated that the knockdown of Mincle in microglia within the PVN prevents VAs by attenuating sympathetic hyperactivity and ventricular susceptibility, in part by inhibiting its downstream NLRP3/IL‐1β axis following MI. Therapeutic interventions targeting Mincle signalling pathway could constitute a novel approach for preventing infarction injury.

// Xia Wang 1 , Junyu Zhao 2 , Yong Zhang 3 , Xiujuan Xue 4 , Jie Yin 3 , Lin Liao 2 , Cuiping Xu 4 , Yinglong Hou 3 , Suhua Yan 3 and Ju Liu 1 1 Medical Research Center, Shandong Provincial Qianfoshan Hospital, Shandong University, Jinan, 250014 Shandong, China 2 Department of Endocrinology, Shandong Provincial Qianfoshan Hospital, Shandong University, Jinan, 250014 Shandong, China 3 Department of Cardiology, Shandong Provincial Qianfoshan Hospital, Shandong University, Jinan, 250014 Shandong, China 4 Department of Nursing, Shandong Provincial Qianfoshan Hospital, Shandong University, Jinan, 250014 Shandong, China Correspondence to: Ju Liu, email: ju.liu@sdu.edu.cn Keywords: von Willebrand factor, kinetics, acute myocardial infarction, meta-analysis Received: May 16, 2017 Accepted: July 26, 2017 Published: August 09, 2017 ABSTRACT Previous studies have shown a variation in plasma level of von Willebrand factor (vWF) in acute myocardial infarction (AMI) patients but with contentious results. In this study, we performed a meta-analysis to evaluate the kinetics of plasma vWF after AMI. A total of 11 qualified studies were obtained through systematical search in PubMed, Web of science, Cochrane Library database and CNKI, followed by search of reference lists, involving 519 AMI patients and 466 non-AMI controls. The standard mean difference (SMD) and 95% confidence intervals (95% CI) were calculated using random-effects model. Results indicated that the plasma vWF was significantly increased in the first several hours after onset of AMI (SMD = 1.94, 95% CI = 1.39–2.48, P 6 h, elevated vWF was found in all 7 days except day 1. Our findings determined the kinetics of plasma vWF after AMI, and might provide a new insight in monitoring AMI progression.

Background Pulmonary arterial hypertension (PAH) is a devastating disease that lacks sufficient treatment. Studies have shown that the Nod-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome contributes to PAH pathogenesis, but the role of the upstream molecular P2X7 receptor (P2X7R) has remained unexplored. We investigated the role of P2X7R in the pathogenesis of PAH. Methods and results PH was induced by a single subcutaneous injection of monocrotaline (MCT) (60 mg/kg) on left pneumonectomised Sprague-Dawley rats, as validated by significant increases in pulmonary artery pressure and vessel wall thickness. Marked P2X7R was detected by predominant PA immunostaining in lungs from PH rats. Western blot revealed a significant increase in the protein levels of P2X7R as well as NLRP3 and caspase-1 in the diseased lung tissue compared with normal tissue. The rats received A-740003 (a selective P2X7 receptor antagonist, 30 mg/kg) daily starting from 1 week before or 2 weeks after MCT injection. Consequently, A-740003 reversed the NLRP3 inflammasome upregulation, significantly decreased the mean right ventricular (RV) pressure and RV hypertrophy, and reversed pulmonary arterial remodelling 4 weeks after MCT injection, as both a pretreatment and rescue intervention. Notably, A-740003 significantly reduced macrophage and pro-inflammatory cytokine levels, as measured via bronchoalveolar lavage. The recruitment of macrophages as well as collagen fibre deposition in the perivascular areas were also reduced, as confirmed by histological staining. Conclusions P2X7R contributes to the pathogenesis of PH, probably in association with activation of the NLRP3 inflammasome. Blockade of P2X7R might be applied as a novel therapeutic approach for the treatment of PAH.

Mounting evidence supports the hypothesis that inflammation modulates sympathetic sprouting after myocardial infarction (MI). The myeloid P2X7 signal has been shown to activate the nucleotide‐binding and oligomerization domain‐like receptor family pyrin domain‐containing 3 (NLRP3) inflammasome, a master regulator of inflammation. We investigated whether P2X7 signal participated in the pathogenesis of sympathetic reinnervation after MI, and whether NLRP3/interleukin‐1β (IL‐1β) axis is involved in the process. We explored the relationship between P2X7 receptor (P2X7R) and IL‐1β in the heart tissue of lipopolysaccharide (LPS)‐primed naive rats. 3′‐O‐(4‐benzoyl) benzoyl adenosine 5′‐triphosphate (BzATP), a P2X7R agonist, induced caspase‐1 activation and mature IL‐1β release, which was further neutralized by a NLRP3 inhibitor (16673‐34‐0). MI was induced by coronary artery ligation. Following infarction, a marked increase in P2X7R was localized within infiltrated macrophages and observed in parallel with an up‐regulation of NLRP3 inflammasome levels and the release of IL‐1β in the left ventricle. The administration of A‐740003 (a P2X7R antagonist) significantly prevented the NLRP3/IL‐1β increase. A‐740003 and/or Anakinra (an IL‐1 receptor antagonist) significantly reduced macrophage infiltration as well as macrophage‐based IL‐1β and NGF (nerve growth factor) production and eventually blunted sympathetic hyperinnervation, as assessed by the immunofluorescence of tyrosine hydroxylase (TH) and growth‐associated protein 43 (GAP 43). Moreover, the use of Anakinra partly attenuated sympathetic sprouting. This indicated that the effect of P2X7 on neural remodelling was mediated at least partially by IL‐1β. The arrhythmia score of programmed electric stimulation was in accordance with the degree of sympathetic hyperinnervation. In vitro studies showed that BzATP up‐regulated secretion of nerve growth factor (NGF) in M1 macrophages via IL‐1β. Together, these data indicate that P2X7R contributes to neural and cardiac remodelling, at least partly mediated by NLRP3/IL‐1β axis. Therapeutic interventions targeting P2X7 signal may be a novel approach to ameliorate arrhythmia following MI.

Background: Inflammation is associated with increased sympathetic drive in cardiovascular diseases. The paraventricular nucleus (PVN) of the hypothalamus is a key regulator of sympathetic nerve activity at post-myocardial infarction (MI). High mobility group box-1 (HMGB1) exhibits inflammatory cytokine like activity in the extracellular space. Inflammation is associated with increased sympathetic drive in cardiovscular diseases. However, the role of HMGB1 in sympathetic nerve activity at post-MI remains unknown. The aim of the present study is to determine the role and mechanism of HMGB1 in the PVN, in terms of sympathetic activity and arrhythmia after MI. Methods: Sprague-Dawley rats underwent left anterior descending coronary artery ligation to induce MI. Anti-HMGB1 polyclonal antibody or control IgG was bilaterally microinjected into the PVN (5 μL every second day for seven consecutive days). Then, renal sympathetic nerve activity (RSNA) was recorded. The association between ventricular arrhythmias (VAs) and MI was evaluated using programmed electrophysiological stimulation. After performing electrophysiological experiments in vivo, immunohistochemistry was used to detect the distribution of HMGB1, while Western blot was used to detect the expression of HMGB1 and p-ERK in the PVN of MI rats. Results: HMGB1 and p-ERK were upregulated in the PVN in rats at post-MI. Moreover, bilateral PVN microinjection of anti-HMGB1 polyclonal antibody reversed the expression of HMGB1 and p-ERK, and consequently decreased the baseline RSNA and inducible VAs, when compared to those in sham rats. Conclusions: These results suggest that MI causes the translocation of HMGB1 in the PVN, which leads to sympathetic overactivation through the ERK1/2 signaling pathway. The bilateral PVN microinjection of anti-HMGB1 antibody can be an effective therapy for MI-induced arrhythmia.

Sympathetic nerve hyperactivity is a primary reason for fatal ventricular arrhythmias (VAs) following myocardial infarction (MI). Pro-inflammatory cytokines produced in the paraventricular nucleus (PVN) post-MI are associated with sympathetic overexcitation; however, the precise mechanism needs further investigation. Our aim was to explore the mechanism of toll-like receptor 4 (TLR4) and its downstream molecular pathway in mediating sympathetic activity post-MI within the PVN. A rat MI model was developed via left anterior descending coronary artery ligation. TLR4 was primarily localized in microglia and increased markedly within the PVN at 3 days in MI rats. Sympathoexcitation also increased, as indicated by high levels of renal sympathetic nerve activity (RSNA) and norepinephrine (NE) concentration. TLR4 knockdown via shRNA microinjection to the PVN resulted in decreased activation of Fos protein (+) neurons in the PVN and peripheral sympathetic nerve activity. TLR4 knockdown also exhibited a lower arrhythmia score following programmed electrical stimulation than those treated with MI surgery only, indicating that the knockdown of TLR4 decreased the incidence of malignant ventricular arrhythmias following MI. LPS-induced inflammatory response was analyzed to explore the underlying mechanism of TLR4 in sympathetic hyperactivity. High levels of NF-κB protein, the pro-inflammatory cytokines IL-1β and TNF-α, and ROS production were observed in the LPS group. PVN-targeted injection of the NF-κB inhibitor PDTC attenuated NF-κB expression and sympathetic activity. Taken together, the results suggested that knockdown of microglial TLR4 within the PVN decreased sympathetic hyperactivity and subsequent VAs post-MI. The downstream NF-κB pathway and ROS production participated in the process. Interventions targeting TLR4 signaling in the PVN may be a novel approach to ameliorate the incidence of VAs post-MI. Keywords: Myocardial infarction, Sympathetic hyperactivity, PVN, TLR4, NF-κB, ROS

Inflammation-dominated sympathetic sprouting adjacent to the necrotic region following myocardial infarction (MI) has been implicated in the etiology of arrhythmias resulting in sudden cardiac death; however, the mechanisms responsible remain to be elucidated. Although being a key immune mediator, the role of Notch has yet to be explored. We investigated whether Notch regulates macrophage responses to inflammation and affects cardiac sympathetic reinnervation in rats undergoing MI. MI was induced by coronary artery ligation. A high level of Notch intracellular domain was observed in the macrophages that infiltrated the infarct area at 3 days post-MI. The administration of the Notch inhibitor N-N-(3,5-difluorophenacetyl-l-alanyl)-S-phenylglycine-t-butyl ester (DAPT) (intravenously 30 min before MI and then daily until death) decreased the number of macrophages and significantly increased the M2 macrophage activation profile in the early stages and attenuated the expression of nerve growth factor (NGF). Eventually, NGF-induced sympathetic hyperinnervation was blunted, as assessed by the immunofluorescence of tyrosine hydroxylase. At 7 days post-MI, the arrhythmia score of programmed electric stimulation in the vehicle-treated infarcted rats was higher than that in rats treated with DAPT. Further deterioration in cardiac function and decreases in the plasma levels of TNF-α and IL-1β were also detected. In vitro studies revealed that LPS/IFN-γ upregulated the surface expression of NGF in M1 macrophages in a Notch-dependent manner. We concluded that Notch inhibition during the acute inflammatory response phase is associated with the downregulation of NGF, probably through a macrophage-dependent pathway, thus preventing the process of sympathetic hyperinnervation.

BACKGROUND This study aimed to decrease leukocytes counts by hydroxyurea (Hu) in an acute myocardial infarction (AMI) rat model and examine its effect on the inflammatory response of myocardial infarction and cardiac functions. MATERIAL AND METHODS AMI was successfully caused in 36 rats, and 12 control rats received sham operation. Rats in the AMI group were then randomly divided into Hu and vehicle group with 18 rats each. Rats in the Hu AMI group received Hu (200 mg/kg) intragastrically while vehicle AMI group received saline. Leukocytes counts, cardiac functions, myocardial tissue morphology, and levels of soluble intercellular adhesion molecule-1 (sICAM), P-selectin and platelet activating factor (PAF) were measured and compared among the three groups four weeks after AMI induction. RESULTS Leukocytes, neutrophils, and leukomonocyte counts in vehicle AMI rats were significantly higher than that of the normal control group (p

Background and objectives Nuclear factor-κB (NF-κB) is considered to be a master inflammation regulator and involved in sympathetic neural hyperinnervation after myocardial infarction (MI). Paraventricular nucleus (PVN), acts as the sympathetic outflow tract, has been proven to play an important role during MI, but whether NF-κB pathway in the PVN participates in the pathogenesis of sympathetic sprouting and reinnervation after MI are unclear. Methods and results MI was induced by coronary artery ligation, NF-κB was activated and interleukin-1β (IL-1β) and nerve growth factor (NGF) levels were increased in the PVN after MI. Lipopolysaccharide (LPS) stimulation also can activate NF-κB pathway, followed by increasing the expressions of IL-1β and NGF in the PVN, once combining with gevokizumab (an IL-1β inhibitor) significantly reduced the expressions of IL-1β and NGF, but could not affect the NF-κB expression in the PVN. Furthermore, micro-injection of the NF-κB inhibitor pyrrolidine dithiocarbamate into PVN in MI rats, significantly inhibited the activation of NF-κB and further reduced the expression of IL-1β and NGF in the PVN, finally blunting sympathetic hyperinnervation in the heart, moreover, the blockade of NF-κB in the PVN reduced the incidence of ventricular arrhythmias (VAs). Conclusions Cardiac nerve sprouting after MI is associated in part with NF-κB activation in the PVN, and IL-1β is involved in the process. Targeting blockade of NF-κB in the PVN may be a potential approach to ameliorate sympathetic hyperinnervation and reduce the incidence of VAs after MI.

Catalpol, an iridiod glucoside isolated from Rehmannia glutinosa, has been reported to possess anti‑inflammatory properties. However, the molecular mechanisms underlying this effect have not been fully elucidated. This study aimed to investigate the effects of catalpol on vascular permeability. Using Transwell permeability assays and measurements of trans‑endothelial electrical resistance (TEER), it was demonstrated that 1 mM catalpol induces a significant increase in the permeability of the monolayers of human umbilical vein endothelial cells (HUVECs). Western blotting and immunofluorescence demonstrated that catalpol inhibits the expression of vascular endothelial (VE)‑cadherin, the key component of adherens junctions, but not occludin, the major constituent of tight junctions. In addition, catalpol inhibits the ETS transcription factor ERG, a positive regulator of VE‑cadherin. Knockdown of ERG expression compromised the catalpol‑induced reduction of TEER in HUVECs. The present study revealed a novel effect of catalpol on vascular permeability and gave insight into the multifaceted roles of catalpol in inflammation.

The von Willebrand factor (vWF) is a plasma glycoprotein that plays an essential role in hemostasis by supporting platelet adhesion and thrombus formation in response to vascular injury. Plasma levels of vWF are an independent risk factor for patients with acute myocardial infarction (AMI); however, clinical data have demonstrated a marked variation of vWF levels in patients with AMI, the reason for which has not yet been identified. In the present study, a rat model of ST-segment elevation AMI was established, and cardiac and peripheral blood was collected for a time-course examination of the plasma levels of vWF and tumor necrosis factor-α (TNF-α). The level of vWF in the blood plasma increased, peaked at 1 h and decreased to normal levels by day 7 following AMI, while the level of TNF-α peaked at 24 h and remained elevated until day 7. The effects of TNF-α on vWF secretion and expression were examined in cultured human umbilical vascular endothelial cells (HUVECs). TNF-α treatment increased vWF secretion from the HUVECs but inhibited the mRNA and protein expression of vWF in the HUVECs. These results indicate that vWF secretion from endothelial cells is transiently elevated following AMI, and then decreases as the expression of vWF is inhibited by TNF-α. The present study increases the understanding of the pathophysiology of vWF and indicates that the determination of vWF levels may be useful in the clinical evaluation of AMI.

The kidney is the principal organ targeted by exposure to cadmium (Cd), a well-known toxic metal. Even at a low level, Cd damages glomerular filtration. However, little is known about the effects of Cd on the glomerular endothelium, which performs the filtration function and directly interacts with Cd in blood plasma. In this study, we cultured human renal glomerular endothelial cells (HRGECs) in the presence of serum with treatment of a short term (1 h) and low concentration (1 μm) of Cd, which mimics the pattern of glomerular endothelium exposure to Cd in vivo. We found that this short-term, low-dose Cd exposure does not induce cytotoxicity, but increases permeability in HRGECs monolayers and redistributes adherens junction proteins vascular endothelial-cadherin and β-catenin. Though short-term, low-dose Cd exposure activates all three major mitogen activated protein kinases, only the inhibitor of p38 mitogen activated protein kinase partially prevents Cd-induced hyperpermeability in HRGECs. Our data indicate that the presence of Cd in blood circulation might directly disrupt the glomerular endothelial cell barrier and contribute to the development of clinical symptoms of glomerular diseases. Copyright © 2015 John Wiley & Sons, Ltd.

Cadmium (Cd) is a persistent and widespread environmental pollutant of continuing worldwide concern. Previous studies have suggested that Cd exposure increases the risk of cardiovascular diseases, such as atherosclerosis and hypertension. However, the underlying mechanisms are poorly understood. In this study, we observed that low dose Cd treatment induced von Willebrand factor (vWF) expression in vascular endothelial cells in mouse lung and kidney tissues. In vitro analysis showed that 1 μM Cd specifically upregulated vWF mRNA and protein expression in human umbilical vein endothelial cells (HUVECs), indicating that Cd targets vascular endothelial cells even at relatively low concentrations. Further study demonstrated that nuclear factor kappa B (NF-κB) and GATA3, two established transcription regulators of the vWF gene, were not altered in the presence of Cd. However, ETS-related gene (ERG) was significantly induced by 1 μM Cd. When ERG was knocked down by siRNA, Cd induced upregulation of vWF was totally blocked. Chromatin immunoprecipitation (ChIP) assay showed that Cd increases the binding of ERG on the −56 ETS motif on the human vWF promoter. These results indicated that ERG mediated the increased expression of vWF by Cd. Since vWF is a key regulator for vascular homeostasis, our findings may provide a novel mechanism for understanding low dose Cd induced development of vascular diseases.

Background: Myocardial infarction (MI) accompanied with abnormal sympathetic innervation, meanwhile, some studies have revealed the oxytocin (OT) and its receptor (OTR) have a relationship with MI and sympathetic system. It is assumed that OT has an close relationship with superior cervical ganglion (SCG), but the existence of oxytocin receptors in SCG has not been well clarified. Objective: Our research aims to explore the expression of OTR in SCG in the setting of MI. Methods and results: MI was induced by coronary artery ligation. Rats were randomly assigned to 2 groups: control, MI. The expression of OTR was measured by Western blotting. Distribution of OTR in SCG was investigated by immunofluorescence. Retrograde tracing test revealed the sprouting of tyrosine hydroxylase (TH: the markers of sensory afferent fibers) from cardiac to SCG neurons. The double-immunofluorescence evidence showed that OTR was co-localized and concomitantly changed with TH and the retrograde neuronal labeling from the cardiac afferent nerves. By Western blotting, the protein of OTR in the MI group was higher than those of the control group. Conclusions: The expression of OTR in SCG after experimental myocardial infarction group was enhanced, suggesting the involvement of OTR in SCG may play a role in the transmission of sympathetic responses after MI.

Inflammation-dominated sympathetic sprouting adjacent to the necrotic region following myocardial infarction (MI) has been implicated in the etiology of arrhythmias resulting in sudden cardiac death; however, the mechanisms responsible remain to be elucidated. Although P2XWe investigated whether P2XAn adenoviral vector with a short hairpin RNA (shRNA) sequence inserted was adopted for the inhibition of P2XThe administration of P2X

Cardiac resynchronization therapy (CRT) is a well-established therapy for patients with heart failure (HF) and wide QRS configuration, especially for those in sinus rhythm. However, for those with permanent AF, atrioventricular nodal (AVN) ablation use remains under debate. Our objective was to evaluate clinical outcomes and mortality of AVN ablation in HF patients with permanent AF receiving CRT. Electronic publication database and reference lists through October 1, 2013 were searched. Observational cohort studies comparing CRT patients with AF who received either AVN ablation or medical therapy were selected. Outcomes included mortality, CRT nonresponse, changes in left ventricular remodeling, and functional outcomes, such as New York Heart Association (NYHA) functional class, quality of life, and 6-minute hall walk distance. Of 1641 reports identified, 13 studies with 1256 patients were included. Among patients with permanent AF and insufficient biventricular pacing (

The vasculature of various organs is a targeted by the environmental toxin, cadmium (Cd). However, mechanisms leading to pathological conditions are poorly understood. In the present study, we examined the effect of cadmium chloride (CdCl2) on human umbilical vein endothelial cells (HUVECs). At 4 μM, CdCl2 induced a hyper-permeability defect in HUVECs, but not the inhibition of cell growth up to 24h. This effect of CdCl2 was dependent on the activation of the p38 mitogen-activated protein kinase (MAPK) pathway. The p38 MAPK inhibitor SB203850 suppressed the CdCl2-induced alteration in trans-endothelial electrical resistance in HUVEC monolayers, a model measurement of vascular endothelial barrier integrity. SB203850 also inhibited the Cd-induced membrane dissociation of vascular endothelial (VE) cadherin and β-catenin, the important components of the adherens junctional complex. In addition, SB203850 reduces the Cd-induced expression and secretion of tumor necrosis factor α (TNF-α). Taken together, our findings suggest that Cd induces vascular hyper-permeability and disruption of endothelial barrier integrity through stimulation of p38 MAPK signaling.

Cardiac sympathetic hyperinnervation after myocardial infarction (MI) is associated with a high incidence of lethal arrhythmia. However, the mechanisms of nerve sprouting induced by MI are unclear. In this study, we showed a nuclear factor-κB (NF-κB) signaling pathway involved in cardiac sympathetic hyperinnervation after MI in rabbit hearts. An MI model was induced by ligation of the coronary artery in rabbits, which were then euthanized after 7 days. Rabbits with MI showed sympathetic hyperinnervation, as revealed by immunohistochemical analysis of the density of nerve fibers positive for growth-associated protein 43 (GAP43) and tyrosine hydroxylase (TH). Using western blot and real-time RT-PCR techniques, we found that MI was associated with activation of NF-κB signaling and consequent upregulation of nerve growth factor. Intravenous administration with the NF-κB inhibitor pyrrolidine dithiocarbamate (100 mg/kg/day) inhibited NF-κB activation and ameliorated sympathetic hyperinnervation after MI. These results suggest that cardiac nerve sprouting after MI is associated in part with NF-κB activation and may be one of the mechanisms responsible for sympathetic hyperinnervation induced by MI.

Background Pulmonary arterial hypertension (PAH) is a devastating disease that lacks sufficient treatment. Studies have shown that the Nod-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome contributes to PAH pathogenesis, but the role of the upstream molecular P2X7 receptor (P2X7R) has remained unexplored. We investigated the role of P2X7R in the pathogenesis of PAH. Methods and results PH was induced by a single subcutaneous injection of monocrotaline (MCT) (60 mg/kg) on left pneumonectomised Sprague-Dawley rats, as validated by significant increases in pulmonary artery pressure and vessel wall thickness. Marked P2X7R was detected by predominant PA immunostaining in lungs from PH rats. Western blot revealed a significant increase in the protein levels of P2X7R as well as NLRP3 and caspase-1 in the diseased lung tissue compared with normal tissue. The rats received A-740003 (a selective P2X7 receptor antagonist, 30 mg/kg) daily starting from 1 week before or 2 weeks after MCT injection. Consequently, A-740003 reversed the NLRP3 inflammasome upregulation, significantly decreased the mean right ventricular (RV) pressure and RV hypertrophy, and reversed pulmonary arterial remodelling 4 weeks after MCT injection, as both a pretreatment and rescue intervention. Notably, A-740003 significantly reduced macrophage and pro-inflammatory cytokine levels, as measured via bronchoalveolar lavage. The recruitment of macrophages as well as collagen fibre deposition in the perivascular areas were also reduced, as confirmed by histological staining. Conclusions P2X7R contributes to the pathogenesis of PH, probably in association with activation of the NLRP3 inflammasome. Blockade of P2X7R might be applied as a novel therapeutic approach for the treatment of PAH. Electronic supplementary material The online version of this article (doi:10.1186/s12931-017-0603-0) contains supplementary material, which is available to authorized users.

Objectives Inflammation after myocardial infarction (MI) causes cardiac nerve sprouting and consequent ventricular arrhythmias (VAs). Macrophages, as major immune cells, are involved in the entire inflammation response process and serve as a link between inflammation and sympathetic hyperinnervation by regulating nerve growth factor (NGF) expression. Accumulating evidence shows that statins possess antiarrhythmogenic properties, and the aim of this study was to explore the mechanism by which atorvastatin ameliorates cardiac sympathetic nerve sprouting via regulating macrophage polarization. Methods Rat models of MI were created by ligating the left coronary artery. MI-operated rats received either atorvastatin or phosphate-buffered saline for 7 days. Immunohistochemical analyses and immunofluorescence staining were used to analyze macrophage infiltration after MI and to detect the distribution and density of growth-associated protein-43 (GAP-43) and tyrosine hydroxylase (TH) in nerve fibers in peri-infarct zones after MI. The polarity of the macrophages that were obtained from the rat peritoneal cavity was examined via flow cytometry. The protein levels of NGF were detected via Western blot analysis, and the concentrations of NGF in the supernatants were determined via enzyme-linked immunosorbent assay. The mRNA levels of NGF, inducible nitric oxide synthase (iNOS), Arginase-1 (Arg1), interleukin-1β (IL-1β), and tumor necrosis factor-α (TNF-α) were examined by quantitative real-time polymerase chain reaction. The association between VAs and MI was evaluated using programmed electrophysiological stimulation. Results Macrophages were successfully induced to the M1 (classically activated) and M2 (alternatively activated) phenotypes by stimulation with lipopolysaccharide and interferon-γ (LPS + IFN-γ) and interleukin-4 (IL-4), respectively. Atorvastatin markedly downregulated IL-1β, TNF-α, iNOS, and NGF and upregulated Arg1, shifting the macrophage phenotype from M1 to M2. Moreover, atorvastatin significantly reduced TH levels and the density of GAP-43-positive nerve fibers and decreased inducible VAs. Conclusion Atorvastatin effectively ameliorated cardiac sympathetic nerve remodeling and prevented VAs after MI by significantly downregulating the expression of NGF, IL-1β, and TNF-α via together with the augmentation of M2 macrophage.

Background To investigate the effects of semaphorin 3A (sema 3A) on cardiac autonomic regulation and subsequent ventricular arrhythmias (VAs) in post-infarcted hearts. Method and results In order to explore the functions of sema 3A in post-infarcted hearts, lentivirus-Sema 3A-shRNA and negative control vectors were delivered to the peri-infarcted myocardium rats respectively. Meanwhile, recombinant sema 3A and control (0.9 % NaCl solution) were injected intravenously into infarcted rats to test the therapeutic potential of sema 3A. Results indicated that levels of sema 3A were higher in post-infarcted hearts compared with sham rats. However, sema 3A silencing leaded to sympathetic hyperinnervation, increased myocardial norepinephrine (NE) content and inducible VAs. Conversely, the intravenous administration of sema 3A to infarcted rats reduced sympathetic nerve sprouting, improved cardiac autonomic regulation, and decreased the incidence of inducible VAs. However, both infarct size and cardiac function were similar among infarcted hearts. Conclusions The upregulation and administration of sema 3A exerted beneficial effects on infarction-induced cardiac autonomic disorders by increasing cardiac electrical stability and reducing VAs. Sema 3A might be a potential therapeutic agent for cardiac autonomic abnormalities induced arrhythmias.

Ce-Al2O3 catalysts prepared by co-precipitation are investigated both in NO oxidation by O2 and in selective catalytic reduction of NO by C2H2 (C2H2-SCR). It is found that C2H2-SCR is initiated and controlled by NO oxidation to NO2 over Al2O3. Ce loading on Al2O3 is almost inactive for NO oxidation below 350°C, since NO2 strongly adsorbs on cerium oxide, leading to the active sites being blocked, which was characterized by temperature-programmed desorption of NO and NO2 and Fourier transform infrared spectroscopy after NO+O2 coadsorption over the samples. However, in the case of C2H2-SCR, Ce loading on Al2O3 significantly improves the reaction by accelerating the NO oxidation step in the temperature range of 250–450°C, since the nitrate species produced by NO2 adsorption is an active intermediate required by C2H2-SCR.

Sixty-three episodes of isthmus-dependent atrial flutter (AFL) in 55 patients were studied to characterize variations in flutter wave morphology and to investigate the mechanisms of the atypical flutter waves on surface ECG. The activation patterns of coronary sinus (CS) and their relationship with flutter wave morphology on the ECG were analyzed. In 46 episodes of counterclockwise AFL (CCW-AFL), there were four types of flutter waves on ECG. Typical and atypical flutter waves were found in 47.8% and 13.0% of the episodes, respectively. Atypical flutter waves had broad positive terminal portion or entirely positive wave in the inferior leads and in V(1), with a distal-to-proximal or fused activation pattern in the CS, and an average activation time of 21.3 +/- 11.4 ms. In 17 episodes of clockwise AFL (CW-AFL), typical and atypical flutter waves were identified in 41.2% and 41.2%, respectively. Atypical flutter waves had negative waves in the inferior ECG leads and in V1, a proximal-to-distal activation pattern in the CS, and an average activation time of 42.4 +/- 14.4 ms. We conclude that atypical flutter waves are common in the isthmus-dependent AFL. The clockwise or counterclockwise conduction in the right atrium, and the activation patterns or conduction sequences between the right and the left atrium, are associated with the variations in the flutter wave morphology on body surface ECG.